Background: Cutaneous leishmaniasis (CL) is a parasitic disease that presents a broad spectrum of clinical features. Treatment of CL is problematic. We aimed to compare the field therapeutic efficacy of topical nanoliposomes containing 0.4% amphotericin B (Nano Lip-AmB) alone and in combination with cryotherapy and/or Glucantime® on human CL in the endemic areas of Iran.

Methods: This retrospective study was performed based on the results of using Nano Lip-AmB alone or with Glucantime® and/or cryotherapy in the treatment of zoonotic cutaneous leishmaniasis (ZCL) in patients referred to health centers of Isfahan, Golestan and Ilam Provinces of Iran as endemic foci of ZCL caused by Leishmania major besides Mashhad and Bam cities as endemic foci of anthroponotic cutaneous leishmaniasis (ACL) caused by with L. tropica.

Results: Two hundred and seventy-eight patients with CL were included in the current study. All of the patients (100%) who received Nano Lip-AmB alone or in combination with Glucantime® and/or cryotherapy based on guideline of Iranian national committee for the treatment of CL. Two patients with 7 skin lesions, who was resident in ACL endemic area and received Nano Lip-AmB plus Glucantime® and another patient was a resident of ZCL endemic area and received Nano Lip-AmB plus cryotherapy showed clinical relapses after treatment.

Conclusion: Sina Ampholeish^® in combination with other standard protocols of treatment of CL is well tolerated and with acceptable clinical efficacy rate.

Background: Echinococcus granulosus is spread by the excretion of cystic organs into the environment. The dog is infected via eating the cystic organ. It then contaminates the environment with eggs of E. granulosus, which are infective to humans and animals. We aimed to determine the E. granulosus genotypes that cause infection in humans in the Van region, Türkiye.

Methods: Sixty patients between 18 and 100 years of age, who underwent the puncture, aspiration, injection, re-aspiration (PAIR) procedure in the Department of Radiodiognastics of Van Yüzüncü Yıl University, Van, Türkiye were included in the study. PAIR fluids were examined microscopically and DNA was isolated from the fluids. After DNA isolation, polymerase chain reaction (PCR) was performed using primers that amplify the E. granulosus NADH dehydrogenase subunit 1 (NAD1) gene region. After sequence analysis of the PCR amplicons, Basic Local Alignment Search Tool (BLAST) was performed.

Results: In the microscopic examination, protoscolex or hook was detected in 42 (70%) of the samples. DNA was successfully extracted from all of the cyst fluids containing protoscolex and hook, and the NAD1 gene region was PCR-amplified. After using BLAST, all of the samples were determined to be an E. granulosus sensu stricto G1 genotype. Sequence comparison revealed that four (9.5%) isolate sequences showed single nucleotide polymorphism (SNP). Sequences of isolates with SNP submitted to the GenBank with accession numbers OR565864 to OR565867. 

Conclusion: E. granulosus s.s. G1 genotype, known as sheep strain, is common in human hydatid disease in the Van region of Türkiye.

Background: Toxoplasma gondii with widespread distribution infects over one third of human populations in the world and can cause serious life-threatening diseases especially for the immunodeficient patients in acute toxoplasmosis. As the clinical pharmaceutical drugs with severe side effects for treatment and non-ideal extant vaccines for prevention, more work starves to be done for keeping advantages in the athletics.

Methods: Aluminum adjuvant and hybrid formaldehyde-killed tachyzoites of T. gondii RH and GT1 isolates were prepared to intramuscularly immunize BALB/c mice for five times at 0, 3, 7, 14 and 21 days post first injection. The triggered humoral and cellular immune responses at two weeks post the last immunization and the survival times of infected mice were examined for the hybrid immunization scheme judgement.

Results: The anti-RH and anti-GT1 specific antibodies were both increased at one week prior to challenge (P < 0.05), and the survival times of immunized mice (7.33 ± 0.71 d for RH, 7.22 ± 0.97 d for GT1) against acute toxoplasmosis were significantly prolonged by the immunizations performed in the study compared to blank control (6.67 ± 0.50 d for RH, 6.33 ± 0.71 d for GT1; P < 0.05), with the higher IFN-γ, IL-2 and IL-12p70 in sera, the elevated CD3e⁺CD4⁺ T and CD3e⁺CD8a⁺ T cells, and the enhanced lymphocyte proliferation in spleen (P < 0.05).

Conclusion: The hybrid killed tachyzoites with aluminum adjuvant induced humoral and cellular immune responses of mice, and offered mildly protective efficacy against acute toxoplasmosis.

Background: Resistance to artemisinin has threatened major achievements in malaria control, more investigations is needed about resistant strains and related genes. We aimed to induce resistance to artesunate in the Plasmodium falciparum 3D7 strain using intermittent exposure method and comparing P.fk13 gene sequence between susceptible and resistance strains.

Methods: P. falciparum 3D7 strain was cultured according to Trager & Jensen method with some modifications. Serial concentrations between 10^-2 mol/l, to 10^-7mol/l were prepared, then P. falciparum 3D7 was exposed to each of the dilution to determine IC50 and lethal dose. Sensitivity reduction process was started from the concentration of 10^-7mol/l and ended at 10^-2mol/l. Exposed parasites were collected after at least 27 days after cultivation in each drug concentration. DNA extraction, PCR and sequencing process were performed to investigate any possible mutations in the P.fk13 gene sequence.

Results: Effectiveness of 10^-2mol/l concentration of artemisinin was found as a lethal dose. IC50 value was equal to 5˟10^-4 mol/l. The resistant strain was provided in the lab, sequenced and registered in the gene bank as P.f Art ^-2, (accession number MH796123. 1). Alignment of this registered sample showed no mutation in P.f kelch13 gene in comparison with standard strain submitted in the GenBank.

Conclusion: Resistance to artesunate in malaria parasite may occur but with no mutation in the P.f kelch13 gene. Therefore, whole genome sequencing should be applied to determine mutations in resistant strains.

Background: We aimed to verify the susceptibility of Leishmania infantum, L. major and L. tropica, to commercial lectins in order to identify the three Leishmania species.

Methods: The degree of agglutination was determined both macroscopically and microscopically and was scored negative (-) to positive (from 1⁺- 4⁺) based on their percentage of agglutination.

Results: Jacalin and UEA-1 were capable of agglutination of L. infantum isolates in both logarithmic and stationary phases at a concentration of 1000 µg/ml (100%). L. tropica isolates showed agglutination with the lectin UEA-1 in both logarithmic and stationary phases (62.5% and 87.5%). L. major and L. tropica showed 75% agglutination with lectin Jacalin in both logarithmic and stationary phases. L. tropica isolates showed 25% agglutination with the lectin WGA in the logarithmic phase. L. infantum, L. major and L. tropica isolates showed 25, 12.5 and 37.5% agglutination in the stationary phase, however, did not show agglutination in logarithmic phases. L. major isolates showed 12.5% agglutination with the lectin PHA in the stationary phase, however, were incapable of agglutination with the L. tropica and L. infantum in both logarithmic and stationary phases.

Conclusion: Despite the fact, that JCA and I-UEA lectins were not able to completely separate L. infantum, L. major and L. tropica. WGA lectin and PHA lectin can help in separating the species of Leishmania parasites.

Background: Sarcocystosis is a zoonotic disease worldwide caused by Sarcocystis spp., some of these species can show clinical and subclinical manifestations, resulting in financial losses. Our study was performed for identifying Sarcocystis sp., in slaughtered buffalo by PCR–RFLP based strategy with sequencing in Guilan, North of Iran.

Methods: Overall, 400 fresh muscle samples were prepared via naked-eye observation from 100 buffaloes (esophagus, diaphragm, shoulder, and thigh), followed by the digestion of samples. The PCR was done to amplify partial parts of the 18S rRNA and mitochondrial cytochrome c oxidase subunit I (Cox1) genes. Then, the PCR products were digested by endonuclease SspI, DraI, and FokI. Sequencing of all species was done to confirm the RFLP results.

Results: Five macroscopic cysts (1.25%) were visible in the sample by naked-eye examination. Furthermore, 293 samples (73.25%) were found to be Sarcocystis sp. positive through tissue digestion and microscopic observation, whereas 376 samples (94%) were positive by PCR. In addition, the findings of PCR-RFLP and nucleotide sequence samples exhibited the infection of buffaloes with S. cruzi.

Conclusion: Based on the data presented herein, Bovine sarcocystosis caused by S. cruzi is very common in buffalo in the Guilan region. Regarding the high prevalence of sarcocystosis, developing disease control and prevention policies for buffaloes is necessary, and a change of attitude in traditional farming is recommended.

Background: Toxoplasmosis could lead to serious outcomes during pregnancy. The aim of this study was to investigate serologic toxoplasmosis in three groups of women according to number of sexual partners.

Methods: The frequency of anti-Toxoplasma IgG from 471 women (101 virgin girls, 240 married women and 130 multi partner women) were determined by ELISA method from referred participant to medical centers of Tehran, Iran in 2020. The results were analyzed by chi-square and logistic regression tests.

Results: Prevalence of toxoplasmosis was significant with the number of sexual partner according to chi square test (P<0.001) and the highest one was observed in multi partners’ women (56.2%) and the lowest one in virgin girls (17.8%). ORs of virgin girls and multi partners’ women were 0.594 and 3.758 respectively, compared to married women. The effect of age on the frequency of anti-Toxoplasma IgG in married women was significant but it was not significant in multi partners’ women. In addition to IgG frequency in married women and multi partners’ women had no significant relationship with the number of children.

Conclusion: Having sexual activity after marriage and having multi partner in sexual activity may possibly be a novel risk factor for toxoplasmosis infection or increasing the IgG frequency.

Background: Canin leishmaniasis (CanL), mostly caused by Leishmania infantum, is one of the most important vector-borne diseases in dogs in the Mediterranean region. In this study, we aimed to determine the disease profile in this region by firstly making microscopic and then molecular analyzes in the samples taken from the dogs.

Methods: Overall, 112 whole blood samples taken from dogs for clinical applications by a veterinarian in Cankırı between December 2021 and November 2022 were used. After blood collection, both thin and thick drop blood smear preparations were prepared and evaluated for Giemsa staining. L. infantum was investigated by Real time-PCR (RT-PCR) method from all blood samples. Sequence analysis and phylogenetic tree study were performed on positive samples.

Results: Both microscopic and RT-PCR analyzes were performed. In both studies, 3 of the 112 samples were positive. Because of the sequence analysis, they were L. infantum. Sequence analysis was performed from the samples found 3 positive. The phylogenetic tree was drawn by making NCBI (National Center for Biotechnology) data entries of the positive samples (Accession numbers: OQ184728, OQ184729, OQ184730).

Conclusion:  Dogs are important, as they are reservoir of this disease. In this study, 3 (2.7%) positive Leishmaniasis was detected in dogs in Cankırı. Ultimately, this should prompt discussion about new strategies going forward to combat infection caused by Leishmania.

Background: We aimed to assess the in vitro effects of the green synthesized silver nanoparticles (Ag NPs) via Thymus vulgaris (thyme) against Leishmania major infection.  

Methods: We have prepared T. vulgalis silver nanoparticles (TSNPs) by adding thyme extract to the silver nitrate aqueous solution (0.2 mM), and evaluated their antileishmanial activity. The viability of L. major promastigotes was assessed in the presence of various concentrations of TSNPs by direct counting after 24 h. The MTT assay was used to identify the viability of promastigotes. The same procedures were assessed in uninfected macrophage cells. The apoptotic effects of nanoparticles on L. major promastigotes were determined by flow cytometry assay using annexin staining. To evaluate anti-amastigotes activity of TSNPs, light microscopic observation was used to determine the number of parasites within the macrophages in each well.

 Results: The effect of TSNPs on promastigotes and amastigotes of L. major was effective and had a reverse relationship with its concentration. TSNPs, inhibited the growth rate of L. major amastigotes and, the IC50 value of these nanoparticles was estimated 3.02 µg/mL (28 µM) after 72h. The results of flow cytometry showed that the toxic effects of TSNPs on promastigotes after 24 hours were statistically significant (P<0.05) and showed 69.51% of apoptosis.

Conclusion: TSNPs had an inhibitor effect on promastigote and amastigote forms of L. major in vitro. It might be considered as a candidate for the treatment of this infection. 

Background: Toxoplasma gondii infects nearly one-third of the world's population. Due to the significant side effects of current treatment options, identifying safe and effective therapies seems crucial. Nanoparticles (NPs) are new promising compounds in treating pathogenic organisms. Currently, no research has investigated the effects of zinc oxide NPs (ZnO-NPs) on Toxoplasma parasite. We aimed to investigate the therapeutic efficacy of ZnO-NPs against tachyzoite forms of T. gondii, RH strain in BALB/c mice.

Methods: In an experiment with 35 female BALB/c mice infected with T. gondii tachyzoites, colloidal ZnO-NPs at concentrations of 10, 20, and 50 ppm, as well as a 50 ppm ZnO solution and a control group, were orally administered four hours after inoculation and continued daily until the mices’ death. Survival rates were calculated and tachyzoite counts were evaluated in the peritoneal fluids of infected mice.

Results: The administration of ZnO-NPs resulted in the reduction of tachyzoite counts in infected mice compared to both the ZnO-treated and control group (P<0.001). Intervention with ZnO-NPs significantly increased the survival time compared to the control group (6.2±0.28 days, P-value <0.05), additionally, the highest dose of ZnO-NPs (50 ppm) showed the highest mice survival time (8.7±0.42 days).

Conclusion: ZnO-NPs were effective in decreasing the number of tachyzoites and increasing mice survival time in vivo. Moreover, there were no significant differences in survival time between the untreated control group and the group treated with zinc oxide, suggesting that, bulk ZnO is not significantly effective in comparison with ZnO-NPs.

Background: Recent studies have shown an increasing number of patients with cutaneous leishmaniasis (CL) who do not respond to pentavalent antimonials as the first line of treatment for CL. Nanocarriers such as extracellular vesicles (EVs) are efficient vehicles that might be used as drug delivery systems for the treatment of diseases. Therefore, we aimed to isolate and characterize the EVs of Leishmania major, load them with Amphotericin B (AmB), and investigate the toxicity and efficacy of the prepared drug form.

Methods: The EVs of L. major were isolated, characterized, and loaded with amphotericin B (AmB), and the EVs-Amphotericin B (EVs-AmB) form was synthesized. Relevant in vitro and in vivo methods were performed to evaluate the toxicity and efficacy of EVs-AmB compared to the control.

Results: The anti-leishmanial activity of the EVs-AmB showed a higher percentage inhibition (PI%) (P = 0.023) compared to the AmB at different concentrations and time points. Obtained data showed a significant increase in the lesion size and parasite load in the lesion, PBS, and EVs mice groups in comparison with EVs-AmB, AmB, and Glucantime groups (P < 0.05), EVs-AmB had a significant decrease in lesion sizes in comparison with AmB (P < 0.05). Results showed that EVs-AmB decreased its toxicity to the kidneys and liver (P < 0.05).

Conclusion: EVs-AmB improved the efficacy of AmB in mouse skin lesions and reduced hepatorenal toxicity. Furthermore, EVs could be a promising nanoplatform for the delivery of AmB in CL caused by L. major.

Introduction: Toxoplasma gondii is an opportunistic protozoan parasite that causes a life-threatening disease – toxoplasmosis – in immunocompromised individuals, including patients with cancer. This prospective cross-sectional study set out to determine the prevalence of toxoplasmosis in patients with cancer compared with that of healthy individuals.

Materials & Methods: Anti-T.gondii IgG and IgM antibodies were measured in the blood samples of 113 patients with cancer (80 with solid organ tumours and 33 with haematological malignancies) and 82 healthy controls, using chemiluminescence microparticle immunoassay (CMIA).

Results: The prevalence of anti-T.gondii IgG was 39.8% in the patient group and 24.4% in the control group, which amounted to a significant difference (p = 0.024). Only one case of anti-T.gondii IgM positivity was observed in the patient group, and no IgM seropositivity was reported in the control group. Moreover, the seroprevalence of anti-T.gondii IgG was non-significantly higher (p = 0.102) in the patients with haematological malignancies (51.5%) than in those with solid organ tumours (35%).

Conclusion: These results demonstrate that the prevalence of T. gondii infection is higher in patients with cancer than in healthy individuals. Therefore, T. gondii screening in patients with cancer is recommended.

Background: The most commonly available drugs for leishmaniasis are pentavalent antimony compounds; whereas the recent studies showed various complications and limitations of these drugs. We aimed to green synthesized silver nanoparticles (AgNPs) and study the promising antileishmanial and synergic effects of green synthesized silver nanoparticles alone and combined with glucantime.

Methods: The precipitation technique was used to drop silver ions via an extract of Astragalus spinosus to AgNPs at Department of Biological Sciences, Faculty of Science and Humanities, Shaqra University, Saudi Arabia in 2022. Then, its anti-amastigotes, caspase-3-like activity, triggering the nitric oxide (NO) as well as its cytotoxicity effects on macrophage cells as well as effects on leishmaniasis in BALB/c mice infected by L. major were measured.

Results: The size of the AgNPs were ranging from 30-40 nm. The IC50 value for AgNPs, AgNPs+ meglumine antimoniate (MA), and MA was 59.3, 18.6, and 51.2 μg/mL, respectively. The determined FIC value for AgNPs and MA was found to be 0.31 and 0.36, respectively; demonstrating the synergistic potency of AgNPs when combined with MA. The diameter of CL lesions treated with various doses of AgNPs and AgNPs+MA notably (p<0.001) decreased. AgNPs, particularly at the concentrations of ½ IC50 and IC50, considerably triggered the caspase-3 activation. The calculated CC50 of AgNPs and MA was 612.5 and 789.8 μg/mL, respectively. Green synthesized AgNPs, especially in combination with MA had synergic antileishmanial effects and displayed a promising drug candidate for treating L. major CL.

Conclusion: We found satisfactory findings in the parasite reduction in both in vitro and animal models. Still, more studies are expected to explain the precise action mechanisms of AgNPs and their efficacy in humans.

Background: Enzymatic digestion of extra cellular matrix proteins by proteinases of   Leishmania promastigotes is a complex process. Hence, studies on functional proteomics of these enzymes can help select these enzymes as possible vaccine candidates or selecting candidates for chemotherapy and immunotherapy. Several proteolytic enzymes are involved in virulence of Leishmania spp. These enzymes are mostly serine, cysteine and metalloproteases. We aimed to detect proteases in Leishmania promastigote exosomes.

Methods: Serine, cysteine and metalloproteases were investigated in exosomes and lysate of L. major promastigote using gelatin zymography. The study was carried out in the Faculty of Veterinary Medicine, University of Tehran, Tehran, Iran, in 2021.

Results: Zymography findings of metalloproteinases showed transparent bands, including a 63-kDa glycoprotein (GP63). This glycoprotein is a major surface metalloproteinase. In addition, transparent bands belonged to serin proteases and cathepsin were demonstrated in gels associated to Leishmania promastigote lysate and exosomes.

Conclusion: Several metalloproteases, serin proteases and cathepsins were shown in promastigote lysate and exosomes of L. major, which could purified and used as fractions for immunodiagnostic.

Background: We aimed to determine species of liver fluke that predominately cause fascioliasis in sheep, goats, and cattle in the Sulaymaniyah Province, Iraq using the molecular technique of DNA sequencing and restriction fragment length polymorphism (RFLP).

Methods: The samples were collected from November 2021 to May 2022. The flukes were collected from infected livers of livestock at the slaughterhouse of Sulaymaniyah Governorate, Iraq. A total of 205 flukes were collected from 56 hosts, cattle (n=22), sheep (n=28), and goats (n=6). The specific primers for FCOX1 and 28S rDNA gene amplification were used. The PCR products were subjected to restriction fragment polymorphism (RFLP) assay using Hpy188III and Dra II restriction enzymes, besides DNA sequencing.

Results: The results showed the genetic polymorphisms among the flukes. Three patterns of RFLP were observed Fasciola hepatica, F. gigantica, and F. intermediate, where 28 of them displayed F. hepatica (sheep, n=14, goat, n=3 and cattle, n= 11), whereas 24 samples displayed the F. gigantica (sheep, n=12, goat, n=3 and cattle, n= 9), and only four samples belonged to F. intermediate (sheep n=3 and cattle, n=1). In addition, the result of the ribosomal DNA (28S rDNA) sequencing confirmed that the isolated flukes belonged to F. hepatica, F. gigantica and F. intermediate.

Conclusion: All three main species are present in the study area and F. hepatica predominated among the animal species in this area also, our results concluded that PCR-RFLP is a rapid and reliable method for liver fluke species identification.

Alveolar echinococcosis (AE) is an important zoonotic tropical disease in China that affects people living in western endemic areas. The disease is prone to occur in the liver with a characteristic similar to slow-growing malignant tumors. We report a 31-year-old male patient with serious complication after hepatorrhaphy, who had presented with clinical manifestations of hepatapostema with infection. Ultrasound (US) and computer tomography (CT) are two important medical imaging modalities to diagnose hepatic AE. Based on the medical history, clinical findings, laboratorial and imaging results, the patient was misdiagnosed with hepatapostema. A series of subsequent treatments were ineffective. Finally, partial hepatectomy was performed, and postoperative pathological results confirmed hepatic AE. The patient has now recovered.

Human alveolar echinococcosis (AE) remains a serious public health concern in endemic areas and a challenge for clinicians. Here a confirmed case of human AE in a patient from Armenia who had not visited a known Echinococcus multilocularis endemic area is reported. In October 2012, a 12-year-old girl from a little Armenian village, presented with paroxysmal pain in the right lumbar area to the children’s medical center (MC). The girl mentioned having close contact with an animal, like a cat. She was admitted to the surgical department with a diagnosis of a malignant liver tumor in the right lobe. In November 2012, the patient underwent laparotomy, removal of the hepatic lesion and abdominal cavity drainage. The histopathological examination of the biopsy material confirmed the main diagnosis of liver AE with suppurative lesions. The patient was given albendazole (ABZ) following 20 days in the hospital, but she stopped receiving the preventive chemotherapy at home and even missed the dispensary visits. It later caused complications, and in July 2016, the child had once again surgery. In January 2017, the child was readmitted to the MC with no content from the external biliary drainage tube in the previous 6 hours. Bile flow improved after flushing the drainage with saline solution and suturing the enterostomy tube. In February 2017, the child visited MC for examination, and the drainage of the bile ducts was blocked, although she had neither discomfort nor jaundice. It was recommended to continue the patient’s follow-up, to receive ABZ and to undergo a liver transplant surgery.